Accelerated osteogenic differentiation of human bone-derived cells in ankylosing spondylitis

Abstract

Ankylosing spondylitis (AS) is characterized by excessive bone formation with syndesmophytes, leading to bony ankylosis. The contribution of osteoblasts to the pathogenesis of ankylosis is poorly understood. The aim of this study was to determine molecular differences between disease controls (Ct) and AS bone-derived cells (BdCs) during osteogenic differentiation with or without inflammation using AS patient serum. We confirmed osteoblastic differentiation of Ct and AS BdCs under osteogenic medium by observing morphological changes and measuring osteoblastic differentiation markers. Osteoblast differentiation was detected by alkaline phosphatase (ALP) staining and activity, and alizarin red and hydroxyapatite staining. Osteoblast-specific markers were analyzed by quantitative reverse-transcriptase-polymerase chain reaction, immunoblotting, and immunostaining. To examine the effects of inflammation, we added AS and healthy control serum to Ct and AS BdCs, and then analyzed osteoblast-specific markers. AS BdCs showed elevated basal intercellular and extracellular ALP activity compared to Ct. When osteoblast differentiation was induced, AS BdCs exhibited higher expression of osteoblast-specific marker genes and faster mineralization than Ct, indicating that these cells differentiated more rapidly into osteoblasts. ALP activity and mineralization accelerated when serum from AS patients was added to Ct and AS BdCs. Our results revealed that AS BdCs showed significantly increased osteoblastic activity and differentiation capacity by regulating osteoblast-specific transcription factors and proteins compared to Ct BdCs. Active inflammation of AS serum accelerated osteoblastic activity. Our study could provide useful basic data for understanding the molecular mechanism of ankylosis in AS.

Regulation of osteoblasts by alkaline phosphatase in ankylosing spondylitis

“…Although blocking agents can reduce inflammation and stabilize the immune system, inhibition of the bony ankylosis process as a therapy remains controversial. Given that high levels of ALP may correlate with disease activity of AS patients, and high osteoblast activity is linked to formation of bony ankylosis in AS, it can be speculated that ALP mediates initiation of osteoblastic differentiation. Thus, it may be further hypothesized that high levels of ALP, through an influence on accelerated osteoblastic activity and differentiation, may lead to ankylosis or new bone formation in AS patients.…”

“…Demographic findings of 10 AS patients are given in Table . Isolation of primary BdCs and osteogenic differentiation methods and activity assessments were carried out as previously reported . Fetal osteoblast cells (FOB, CRL‐11372) were purchased from the American Type Culture Collection (ATCC) and cultured in Dulbecco's modified Eagle's mudium (DMEM):F12 (Hyclone, South Logan, UT, USA, SH30025.01).…”

“…Ankylosing spondylitis (AS) is an inflammatory bone disease, characterized by excessive and accelerated osteoblastic activity . The disease initially presents as inflammation, followed by altered osteoblastic activity that can result in bony ankylosis .…”

HLA-B27–mediated activation of TNAP phosphatase promotes pathogenic syndesmophyte formation in ankylosing spondylitis

“…The lack of treatments is associated with our incomplete understanding of the causative pathological roles of stromal activation, lack of a proper platform for drug screening, and absence of good biomarkers to select a high-risk group of patients who are most likely to exhibit radiographic progression. Although the accelerated osteogenesis of AS patient-derived bone marrow (BM) mesenchymal stem cells (MSCs) from hips or bonederived precursors from facet joints has been reported (12)(13)(14)(15), the role of MSCs from the enthesis or nidus of syndesmophytes, and the underlying genetic pathways involved, are largely unknown. Herein, we isolated MSCs from the enthesis involved in the spinal ankylosis of AS patients (AS MSCs) and found that the genetic HLA-B27-mediated spliced X-box-binding protein 1 (sXBP1)/ retinoic acid receptor-β (RARB)/tissue-nonspecific alkaline phosphatase (TNAP) pathway contributed to the syndesmophyte formation of AS.…”

Loss and gain of bone in spondyloarthritis: what drives these opposing clinical features?

“…Furthermore studies have shown that cells derived from the facet joints of AS patients, as opposed to cells from spinal injury due to trauma, have an increased osteogenic capacity. 77 Thus, the combination of reduced Wnt inhibition, altered mechanical strain, and an increased propensity to form osteoblasts could all contribute to syndesmophytes formation in axSpA.…”